Analysis of tocopherols by gas-liquid and high-performance liquid chromatography: a comparative study

Abstract

Analyse von Tocopherolen mittels Gas-Flüssigkeits-und Hochleistungsflüssigchromatographie: eine vergleichende Untersuchung. Ausgewählte Normalphasen-HPLC-und gaschromatographische Methoden wurden für die Analyse von Tocopherolen in pflanzlichen Ölen untersucht. Wiederholbarkeit, Genauigkeit und/oder Wiederfindung wurden für beide Methoden bestimmt und mit Daten der offiziellen Methodik verglichen. Die größte Genauigkeit wurde mittels der Normalphasen-HPLC-Technik, basierend auf der AOCS-Methode Ce 8-89, erzielt, was wahrscheinlich durch die kurze und einfache Probenaufbereitung erklärt werden kann. Es wurden Tocopherolgehalte von sechzehn voll raffinierten Pflanzenölen mit sowohl GLC als auch HPLC bestimmt. Die Regressionsanalyse zeigte, daß Tocopherolgehalte, die mittels GLC ermittelt worden sind, um 6,1% höher waren als die vergleichbaren HPLC-Daten. Obwohl die ermittelte Wiederfindungsrate der GLC-Methode gut ist, scheint es, daß bei der Probenaufbereitung zusätzlich eine Auftrennung mittels Dünnschichtchromatographie eingeführt werden sollte, um eine Überlappung der Peaks zwischen Tocopherolen und einigen störenden Substanzen der unverseifbaren Fraktion zu vermeiden.a Department of Food Technology and Nutrition, University of Ghent, Gent, Belgium. b De Smet Engineering N. V., Edegem, Belgium.vents and reagents used were analytical or HPLC grade and purchased from Acros Organics (Geel, Belgium). Gas liquid chromatographySample preparation: The selected GLC procedure was based on the AOCS official method Ce 3-74 [6]. 5-7 g of oil were saponified in the presence of 0.15 % butylhydroxytoluene as antioxidant [5]. Tocopherols and sterols were not separated by thin-layer chromatography prior to GLC analysis. Derivatization was accomplished by the addition of 1 ml acetic acid anhydride, keeping the mixture 10 min at 140 °C. After cooling, 1 µl was injected in the gas chromatograph.Gas-chromatographic conditions: Analyses were carried out on a Carlo Erba 4160 chromatograph equipped with a flame-ionization detector (FID) and a split injector (split ratio 1:40) (Carlo Erba, Milano, Italy). A fused silica capillary column coated with a nonpolar stationary phase (CP TM Sil 5CB, 25 m × 0.25 mm internal diameter; 0.25 µm film thickness, Chrompack, The Netherlands) was selected. The oven temperature was kept constant at 290 °C. Injector and detector were maintained at 340 °C. All tocopherol homologues eluted within 15 min. The total analysis time was set at 30 min to assure the elution of all sterols.Identification of the individual tocopherols was realized with pure standards as reference compounds.Quantification: Correction factors were determined for all tocopherols by analyzing different mixtures of the tocopherol standards and an internal standard (3-octadecyloxy-1,2-propanediol). Quantitative analyses were performed with a Shimadzu RC3A integrator. High-performance liquid chromatographyPrinciple: The selected NP-HPLC procedure was based on the previously mentioned IUPAC method 2.432 [5] and the AOCS official method Ce ...

Influence of chemical refining on the major and minor components of rice brain oil

“…The level of γ-oryzanol was determined spectrophotometrically according to the method described by Seetharamaiah and Prabhakar (15). Tocopherol and tocotrienol contents were analyzed by normalphase HPLC (Alltima Silica 5U, 250 mm × 4.6 mm i.d., particle size 5 µm; Alltech, Lokeren, Belgium) based on AOCS Official Method Ce 8-89 (16).…”

Antioxidant activity screening of extracts from Sideritis species (Labiatae) grown in Bulgaria

“…It contained 562 ppm α-tocopherol and traces of γ -tocopherol as determined by high-performance liquid chromatography (HPLC). 26 The oil was stored under nitrogen in the dark at −20 • C. Fatty acid composition determined by GC as methyl esters was as follows: 6.9% C16:0, 2.7% C18:0, 12.6% C18:1, 77.5% C18:2. All solvents used were of analytical grade or HPLC grade (for HPLC analyses).…”

A rapid in‐house validated GC‐FID method for simultaneous determination of lipophilic bioactives in olive oil: Squalene, α‐tocopherol, and β‐sitosterol

“…Silylation (simple reactions that involve replacement of active hydrogen (OH, COOH) by a silyl group reducing hydrogen bonding and the polarity of the compound), which is one of the most prevalent derivatization methods , is commonly used for determination of sterols. It is also preferred for tocopherol determination rather than alkylation and acylation, which contain difficult esterification reactions and procedures (heating at 100–140°C) .…”

“…Silylation (simple reactions that involve replacement of active hydrogen (-OH, -COOH) by a silyl group reducing hydrogen bonding and the polarity of the compound), which is one of the most prevalent derivatization methods [37], is commonly used for determination of sterols. It is also preferred for tocopherol determination rather than alkylation and acylation, which contain difficult esterification reactions and procedures (heating at 100-140°C) [38]. The silylation mixture, hexamethyldisilazane (HMDS, a weak silylating reagent): TMCS (catalyst): pyridine (polar solvent), was added (260 mL, 3:1:9, v/v) to dry unsaponifiable matter of olive oil (<1%) [10,26].…”